The objective of our research is to examine the role of various gene products of bacteriophage T4 in DNA replication, recombination and repair. Special emphasis will be given to the DNA-unwinding protein (gene 32 product, gp32) which is not only a key component in reactions mentioned above, but also regulates many other enzymes (DNA Ligase, DNA polymerase and nuclease) through direct interactions. Previously, we purified three stable proteolysis intermediates of gp32 and started comparative physicochemical studies of some of their properties such as affinities for DNAs, DNA-unwinding profiles, circular dichroism and electronmicroscopy of their DNA-complexes. From results of these studies, we proposed the following: The NH2-terminal peptide (10-15 amino acids) is essential for tight binding to single-stranded DNA and protrudes from the native free gp32 molecule but is buried inside the protein DNA-complex. The COOH-terminal (30-50 amino acids) protrudes from either the free- or DNA-bound molecule and has a regulatory role inhibiting DNA melting activity. Its reversible displacement through interaction of other protein components within the replication apparatus or its proteolytic removal in vitro turns gp32 from a DNA-renaturing to a DNA-denaturing protein. The protruded part of the molecule is probably also related to regulation of other enzyme activities. We plan to continue our comparative physico-chemical examinations of gp32 and its modified forms (proteolysis intermediates, ts-proteins, amber peptides) to elucidate DNA-binding, unwinding and renaturation mechanisms. We also propose studying their effects on other enzyme activities and their activities in in vitro DNA-replication and recombination systems. We will extend proteolysis experiments to DNA-unwinding proteins from other origins.